Abstract   The Cache Slough complex is an area of tidal sloughs in the Sacramento–San Joaquin River Delta of California (USA), and is surrounding by irrigated agricultural lands. Among the species of concern in the area is the delta smelt (Hypomesus transpacificus), a federally listed threatened species. Releases of the organophosphate insecticide chlorpyrifos and pyrethroid insecticides were examined to determine whether they represented a threat to the copepods on which delta smelt feed (Eurytemora affinis and Pseudodiaptomus forbesi) and to aquatic life in general, represented by the standard testing organism, Hyalella azteca. There was a single incident of toxicity to H. azteca as a result of discharge of agricultural irrigation water containing chlorpyrifos. Pyrethroids were not found in samples collected during the dry season. Following rain events, however, the waters of western Cache Slough repeatedly became toxic to H. azteca because of the pyrethroids bifenthrin and cyhalothrin. The 96-h median lethal concentrations (LC50s) for E. affinis and P. forbesi for the pyrethroids bifenthrin and cyhalothrin were 16.7 ng/L to 19.4 ng/L when tested at 20 °C. However, their LC50s may be 5 mg/L to 10 ng/L at in situ temperatures of the Cache Slough, comparable to the peak bifenthrin concentration observed. The dominant pyrethroid source appeared to be urban runoff entering a creek 21 km upstream of Cache Slough. Pyrethroids of urban origin were supplemented by agricultural inputs of pyrethroids and chlorpyrifos as the creek flowed toward Cache Slough. Environ Toxicol Chem 2014;33:920–929. © 2014 SETAC